Arabinofuranosidases are important enzymes in the liberation of arabinose from xylan and pectin. In the genome of Aspergillus fumigatus, two genes encoding putative GH family 62 arabinofuranosidases (abf1 and 2) have been identified. The sequence of abf2 is 1191 bp in length including an intron and a carbohydrate binding module (CBM), while abf1  with 999 bp shows no introns and CBMs. When the catalytic modules of the respective proteins are compared, they show a 79% identity. The estimated molecular masses of the proteins are 36428 and 42950 for ABF1 and 2, respectively. Using Pichia pastoris as host, both proteins have been expressed heterologously and purified from the culture supernatants. ABF1 has a molecular mass of about 30 KDa and ABF2 of 43 KDa as estimated from SDS gels. Both enzymes are active using p-nitrophenyl arabinofuranoside (pNPAra) as substrate, and show a pH optimum of 4-5 (ABF1) and 5 (ABF2). Optimal temperatures are 37�C and 42 �C, respectively for ABF 1 and 2. Km for ABF1 is 94.2 mM and for ABF2 3.9 mM. ABF2 is considerably more stable: at 45�C and 20 min incubation it retains 80% of its activity, compared to 20% for ABF1. Both enzymes liberate arabinose from arabinoxylan and show little activity towards arabinan or debranched arabinan in agreement with other fungal family 62 ABFs. Although both enzymes show similarities in substrate specificity, the presence of a CBM in ABF2 increases its affinity for pNPAra and its heat stabiliby.Grant support: FONDECYT 1130180 and UNAB DI-61-12/R

Aspergillus fumigatus produces two arabinofuranosidases belonging to glycosyl hydrolase family 62, one of them possessing a carbohydrate binding module: Heterologous expression and characterization of both enzymes

248th National Meeting (2014)

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